AIDS is caused by HIV infection of human cells, primarily T4 lymphocytes. The recognition process of the virus by the human cells is mediated by the viral envelope glycoprotein gp120 and the cellular receptor glycoprotein for gp120, known as CD4. Analogous to many other cell-cell and cell-molecule interactions, the carbohydrate portions of gp120 and CD4, exposed as they are to the outer surface of the virus particle and the T4 cells, respectively, are likely to play a key role in the binding of HIV to CD4. To understand their functional involvement in the interaction of HIV-1 with T4 lymphocytes, knowledge of the structure of the carbohydrate moieties of these cell-surface glycoproteins is a prerequisite. Specifically, our research is aimed at characterizing the carbohydrate side chains of recombinantly expressed analogs of the HIV surface glycoprotein gp120. The latter are (constituents of) candidate prophylactic anti-AIDS vaccines currently in clinical trials. The reason for the attention to these immunogens in vaccine development is the opinion that antibodies directed against the viral particle surface is most likely to mediate the desired protective effect. We have completed our structural studies of the carbohydrates of recombinant envelope glycoprotein gp120 of HIV type 1, strain IIIB, expressed in Chinese hamster ovary (CHO) cells provided to us by Dr. Spellman. The CHO cell-derived rgp120 is a highly glycosylated protein, containing 24 occupied N-glycosylation sites. All 24 sites have been characterized, with 11 sites containing high-mannose structures, 11 sites the complex-type, and 2 sites containing mixtures of all three types. The carbohydrate portions of the glycoprotein play an important role in all the structural and functional features of the molecule, such as structural variability and heterogeneity, three-dimensional structure, antigenicity, and the biologically important recognition and binding to the cell-surface receptor CD4. Detailed knowledge of the carbohydrate structures of rgp120, in particular those close to the V3 domain of the protein, should lead to an improved understanding of their function in the recognition and binding of HIV to its host's cells. Our integrated structural analytical approach involved a combination of enzymatic digestions of the rgp120, followed by extensive chromatographic isolation and purification of oligosaccharides and glycopeptides. The latter were characterized by a combination of 1H-NMR spectroscopy, MALDI-TOF and electrospray mass spectrometry, Dionex HPAEC, capillary electrophoresis, and FT-IR spectroscopy.